Disruption of SorCS2 reveals differences in the regulation of stereociliary bundle formation between hair cell types in the inner ear.
Disruption of SorCS2 reveals differences in the regulation of stereociliary bundle formation between hair cell types in the inner ear.
复制标题
DOI：
10.1371/journal.pgen.1006692
复制
发表时间：
2017-03
期刊：
影响因子：
4.5
通讯作者：
Jagger DJ
中科院分区：
文献类型：
--
作者：
Forge A;Taylor RR;Dawson SJ;Lovett M;Jagger DJ
在一群转基因小鼠中观察到暗示内耳疾病的行为异常。受影响的动物严重失聪。严重的毛束缺陷被确定在所有外毛细胞和内毛细胞（OHC，IHC）在耳蜗和前庭黄斑器官的毛细胞，但在嵴毛细胞基本上不受影响。有证据表明，这种疾病可能是由于随机插入的转基因构建体造成的基因破坏。全基因组测序鉴定了SorCS 2（Sortilin相关VPS-10结构域蛋白）基因座的中断。实时-qPCR显示受影响小鼠耳蜗组织中SorCS 2 RNA的表达被破坏，这通过SorCS 2免疫标记得到证实。在所有受影响的毛细胞，静纤毛短于正常，但异常的束形态和组织不同的毛细胞类型。OHC上的束严重畸形，静纤毛明显少于正常。然而，静纤毛组织在行的高度增加。IHC上的束状体包含比正常明显更多的静纤毛，其中一些更长的静纤毛朝向中心，或具有最小的高度差。在出生后早期的小鼠，动纤毛（初级纤毛）的IHC和OHC最初位于朝向毛细胞表面的外侧边缘，但往往成为包围静纤毛束形状和顶面轮廓的变化。在黄斑器官中，在整个成熟过程中，动纤毛位于细胞表面的中心。有中断的信号通路控制内在的毛细胞顶端不对称。LGN和Gαi3大部分缺失，非典型蛋白激酶C（aPKC）失去其不对称分布。结果表明，SorCS 2在内在极性通路的上游发挥作用，并且毛细胞类型之间在产生精确组织的毛束的机制的部署方面存在差异。内耳中的感觉“毛”细胞的名称来源于其顶表面上的机械感觉“静纤毛”的组织束。指状静纤毛由肌动蛋白丝组成，在细胞表面的一个方向上呈阶梯状增加，这是一种在感觉上皮上提供明确极性的系统。毛束极性和方向的建立是一个涉及多个信号通路的高度调节的过程。毛细胞亚型之间的束状形态存在细微差异，表明这些调控途径存在局部差异。在一个转基因小鼠群体中，我们发现严重失聪的动物在所有听觉毛细胞上都表现出严重的毛束异常。平衡系统重力感受组织中的毛细胞也有异常束，但对感受旋转的毛细胞束没有影响。毛束异常与内源性顶端不对称途径蛋白质的错误表达相一致。这些缺陷与SorCS 2基因的破坏以及随后内耳中编码的跨膜受体的错误表达有关。结果表明，SorCS 2调节毛细胞发育，并且毛细胞亚型形成其静纤毛束的方式存在以前未被认识到的差异。
Behavioural anomalies suggesting an inner ear disorder were observed in a colony of transgenic mice. Affected animals were profoundly deaf. Severe hair bundle defects were identified in all outer and inner hair cells (OHC, IHC) in the cochlea and in hair cells of vestibular macular organs, but hair cells in cristae were essentially unaffected. Evidence suggested the disorder was likely due to gene disruption by a randomly inserted transgene construct. Whole-genome sequencing identified interruption of the SorCS2 (Sortilin-related VPS-10 domain containing protein) locus. Real-time-qPCR demonstrated disrupted expression of SorCS2 RNA in cochlear tissue from affected mice and this was confirmed by SorCS2 immuno-labelling. In all affected hair cells, stereocilia were shorter than normal, but abnormalities of bundle morphology and organisation differed between hair cell types. Bundles on OHC were grossly misshapen with significantly fewer stereocilia than normal. However, stereocilia were organised in rows of increasing height. Bundles on IHC contained significantly more stereocilia than normal with some longer stereocilia towards the centre, or with minimal height differentials. In early postnatal mice, kinocilia (primary cilia) of IHC and of OHC were initially located towards the lateral edge of the hair cell surface but often became surrounded by stereocilia as bundle shape and apical surface contour changed. In macular organs the kinocilium was positioned in the centre of the cell surface throughout maturation. There was disruption of the signalling pathway controlling intrinsic hair cell apical asymmetry. LGN and Gαi3 were largely absent, and atypical Protein Kinase C (aPKC) lost its asymmetric distribution. The results suggest that SorCS2 plays a role upstream of the intrinsic polarity pathway and that there are differences between hair cell types in the deployment of the machinery that generates a precisely organised hair bundle. Sensory “hair” cells in the inner ear derive their name from an organised bundle of mechano-sensory “stereocilia” on their apical surface. The finger-like stereocilia are composed of actin filaments and increase in height staircase-like in one direction across the cell surface, a system which provides a defined polarity across the sensory epithelium. The establishment of hair bundle polarity and orientation is a highly regulated process involving several signalling pathways. There are subtle differences in bundle morphology between hair cell subtypes, suggesting local variations exist within these regulatory pathways. In a transgenic mouse colony we found profoundly deaf animals displaying severe hair bundle abnormalities on all auditory hair cells. Hair cells in the gravity sensing tissues of the balance system also had abnormal bundles, but there was no effect on bundles of hair cells sensing rotation. Hair bundle abnormalities coincided with mis- expression of proteins of an intrinsic apical asymmetry pathway. These defects were associated with disruption of the SorCS2 gene, and consequent mis-expression of the coded transmembrane receptor in the inner ear. The results suggest that SorCS2 regulates hair cell development, and that there are previously unrecognised differences in the way subtypes of hair cells form their stereociliary bundles.